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PDBsum entry 2z2e
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References listed in PDB file
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Key reference
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Title
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Spontaneous asparaginyl deamidation of canine milk lysozyme under mild conditions.
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Authors
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Y.Nonaka,
T.Aizawa,
D.Akieda,
M.Yasui,
M.Watanabe,
N.Watanabe,
I.Tanaka,
M.Kamiya,
M.Mizuguchi,
M.Demura,
K.Kawano.
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Ref.
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Proteins, 2008,
72,
313-322.
[DOI no: ]
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PubMed id
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Abstract
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Asparaginyl deamidation is a common form of nonenzymatic degradation of proteins
and peptides. As it introduces a negative charge spontaneously and irreversibly,
charge heterogeneity can be accumulated in protein solution during purification,
preservation, and experiments. In this study, canine milk lysozyme (CML), a
useful model for the study of the molten globule state, exhibited charge
heterogeneity after sample purification. Four Asn residues in CML deamidated
rapidly under mild conditions: pH 8.0 and 30 degrees C. Other than these
residues, one Asn residue, which was stable in the native state, was labile to
deamidation in the unfolded state. This suggests that the structural formation
around Asn can suppress deamidation. Substitutions of these labile Asn residues
to Gln residues prevented deamidation effectively. Because the substitutions did
not disrupt the structural formation of the native and molten globule states,
they will enable more precise analyses for physical and structural studies.
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Figure 1.
Figure 1. The currently accepted mechanism of asparagine
nonenzymatic deamidation.[11][12] The succinimide intermediate
formation arises from a nucleophilic attack of the backbone NH
group of the next residue on the side-chain carbonyl of the
asparaginyl residue. The ring is hydrolyzed to open, resulting
in either an aspartic acid or isoaspartic acid product.
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Figure 6.
Figure 6. (a) Crystal structure of 5NQ. The substituted
residues are represented in yellow. (b) Superposition of the
backbone structures of CML wild type (red, PDB 2CWI) and 5NQ
(blue) by X-ray crystallography. The root-mean-square deviation
of the backbones of the two structures is 0.511 Å.
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The above figures are
reprinted
by permission from John Wiley & Sons, Inc.:
Proteins
(2008,
72,
313-322)
copyright 2008.
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